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Matilda F. Chan, Jing Li, Anthony A. Bertrand, Inna A. Maltseva, Zena Werb; Role of Macrophage Elastase (MMP12) in Corneal Myeloid Cell Activities Visualized With Time-lapse in vivo Confocal Microscopy. Invest. Ophthalmol. Vis. Sci. 2011;52(14):1123.
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To analyze myeloid cell activity in response to corneal chemical injury and to investigate the role of macrophage elastase (MMP12).
Corneal expression of MMP12 in wild-type (WT) mice following chemical injury was assayed using quantitative real-time PCR (qPCR). Corneal macrophage infiltration of injured WT and MMP12-/- mice was quantified using whole-mount immunofluorescence staining (F4/80 antibody) and flow cytometry. Relative expression levels of CC chemokines were compared using qPCR and protein arrays. Mice expressing EGFP under the c-fms promoter were crossed with the MMP12 line to generate WT and MMP12-/- mice expressing EGFP-labeled myeloid cells to visualize these cells in the corneal stroma. Myeloid cell activity was documented in vivo with time-lapse confocal microscopy (20-90 minute duration) before and 1 day after injury. Imaging software (Imaris) was used to create movies tracking cell movement.
MMP12 expression levels following corneal chemical injury of WT mice showed peak expression levels occurring 2 and 6 days after injury. Injured corneas of MMP12-/- mice demonstrated increased macrophage infiltration compared with WT mice. Members of the CC chemokine family were screened and higher levels of monocyte chemoattractant protein-1 (MCP-1 or CCL2) were detected in wounded corneas of MMP12-/- mice. In vivo time-lapse confocal microscopy allowed visualization and comparison of myeloid cell behavior in WT and MMP12-/- mice. Resident corneal myeloid cells of non-injured WT and MMP12-/- mice displayed minimal movement. One day after chemical injury, myeloid cells of WT mice were active and had increased track length. In contrast, myeloid cells of injured MMP12-/- mouse corneas remained minimally active and tracking plots showed little migration.
MMP12 deficient mice had elevated CCL2 expression and increased corneal macrophage accumulation in response to chemical injury suggesting a role for MMP12 in the regulation and distribution of myeloid cells. We were able to monitor in vivo myeloid cell activities using time-lapse confocal microscopy. Upon injury, myeloid cells in WT mice became more migratory likely representing a scavenger response. Myeloid cells of MMP12-/- mice, however, remained minimally migratory which may reflect a difference in macrophage M1 and M2 polarization. These findings suggest an important role for MMP12 in the regulation of myeloid cell activities in the corneal healing response.
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